Project Summary Intact proteins and multi-protein complexes represent two of the most important levels of organization in biology, yet these levels are also some of the most difficult to study. Top-down proteomics offers a bold vision: direct observation, identification, and relative quantification of all the protein forms in a biological sample. Native mass spectrometry aims at the next higher level of organization by providing a means to observe proteins in complex with ligands or other proteins. The proposed project will build new algorithms and software for intact, native, and top-down mass spectrometry. Specifically, the project will further develop Protein Metrics Intact Mass charge- deconvolution software, incorporating multiple information channels (isotope spacing, co-elution, and charge states) into the core algorithm, so that the algorithm can handle complex, low signal- to-noise mass spectra over a wide range of masses and abundances. The project will also build differential quantification (?proteoform profiling?) for undigested proteins and complexes, regardless of whether or not the molecules have isotope resolution. The project will enable a number of interesting applications, including biomarker discovery, large-molecule drug comparability, pharmacokinetics, and protein / ligand and protein / protein binding.